The lack of an absolutely effective paradigm to curb human immunodeficiency virus type-1 or type-2 (HIV-1/-2) infection warrants identification of novel diagnostic and therapeutic products. We have developed a rapid and sensitive assay to measure the cell associated virus in peripheral blood cells from HIV-infected subjects. 9NC, an inhibitor of cellular topo-I enzyme, blocks HIV-1 replication in vitro in a latently infected human T-cell clone. Virus replication was initially activated by various inducers such as the cytokine tumor necrosis factor-alpha (TNF) which replaces the functional HIV-1 Tat protein and appears to promote disease progression. Both HIV-1 and HIV-2 tat-defective proviruses can efficiently be rescued by TNF. We found about 80% inhibition of HIV-1 RNA and a consistent reduction of virus in culture up to about 95% when the cells were exposed to 9NC. However, 9NC failed to suppress a similar virus activation mediated by sodium butyrate. Our data suggest that 9NC is a potent drug in the suppression of NF-kB-mediated, but not histone-acetylation-mediated pathway of virus activation. 9NC could prove to be an effective therapeutic drug in maintaining a low level of viremia by suppressing the activation of latent proviral DNAs by cytokines such as TNF, while attenuating the neoplastic changes in AIDS-associated malignancies.